Convenient syntheses of 2-acylamino-4-halothiazoles and acylated derivatives using a versatile Boc-intermediate

The 2-aminothiazole grouping is a significant feature of many series of biologically active molecules, including antibiotics, anticancer agents and NSAIDs. We have a longstanding interest in the synthesis and biological evaluation of thiazolides, viz. [2-hydroxyaroyl-N-(thiazol-2-yl)-amides] which have broad spectrum antiinfective, especially antiviral, properties. However, 2-amino-4-substituted thiazoles, especially 4-halo examples, are not easily available. We now report practical, efficient syntheses of this class from readily available pseudothiohydantoin, or 2-aminothiazol-4(5H)-one: the key intermediate was its Boc derivative, from which, under Appel-related conditions, Br, Cl and I could all be introduced at C(4). Whereas 2-amino-4-Br/4-Cl thiazoles gave low yields of mixed products on acylation, including a bis-acyl product, further acylation of the Boc intermediates, with a final mild deprotection step, afforded the desired thiazolides cleanly and in good yields. In contrast, even mild hydrolysis of 2-acetamido-4-chlorothiazole led to decomposition with fast reversion to 2-aminothiazol-4(5H)-one. We also present a correction of a claimed synthesis of 2-acetamido-4-chlorothiazole, which in fact produces its 5-chloro isomer.


Introduction
A 2-aminothiazole unit is a common feature of many biologically active molecular series, 1 such as cephalosporin antibiotics, kinase inhibitor anticancer agents and non-steroidal antiin-ammatory drugs, Fig. 1.It has been suggested that 2-aminothiazole substitution favourably affects both the activity prole and absorption properties. 2,3Although a thiazole unsubstituted at both C(4) and C( 5) is regarded as a metabolic risk, 4 this danger is readily averted by appropriate substitution, especially with electron-withdrawing substituents. 5ne important class of broad spectrum antiinfective 2-aminothiazole derivatives are the thiazolides, or [2-hydroxyaroyl-N-(thiazol-2-yl)-amides], typied by nitazoxanide 1a which was rst reported in 1975 (Fig. 2). 6To this day 1a remains the antiparasitic agent of choice against Cryptosporidium spp. 7It was later discovered that 1a and other analogues, notably the 5chloro analogue 1b, were broad-spectrum antiviral agents, [8][9][10] dating from the use of 1a in treating cryptosporidiosis in AIDS patients.
We have described the structure-activity relationships (SAR) of a wide range of thiazolides against hepatitis B, hepatitis C and inuenza A viruses. [11][12][13] Against a typical H1N1 strain of inuenza A virus, compound 1a shows IC 50 = 3.3 mM and 1b shows IC 50 = 3.4 mM.13b Clinical trials of 1a have been performed against rotavirus 14 and acute uncomplicated inuenza A. 15a,b More recently, the SARS-CoV2 pandemic led to a strong resurgence of interest in small molecule antivirals, and NTZ has shown notable activity in trials against SARS-CoV2. 16The active circulating metabolites of 1a/1b in vivo are the free phenols 2a/ 2b, of which the phenolic acetates are prodrugs. 17Later we prepared more efficient, amino-acid ester prodrugs 3a/3b, which were shown to offer greatly improved bioavailability compared to 1a/1b. 18n general, 5-substituted thiazolides such as 1a/1b are the easiest to obtain.The natural position of electrophilic substitution of a 2-aminothiazole is at position 5, even when the 2amine is acylated.In order to synthesise thiazolides with a 4substituent, including 4-halo examples, various methods are possible: the 4-sulfonyl thiazolide 4 was synthesised from a thioester.13b One approach to a 2-amino-4-bromothiazole uses the halogen dance rearrangement from a protected 5-Br thiazole, as originally described by Stangeland and Stanetty (Scheme 1), 19,20 employing LiNPr 2 i in THF.The rearrangement of 5 to 6 is considered to proceed via the N, C(5)-dianion which is thermodynamically preferred (Scheme 1, lower).This proved a robust procedure, but on removal of the Boc group the free amine 7, Fig. 3, proved rather unstable and difficult to acylate, in contrast to 2-amino-5-bromothiazole.The literature on 2-amino-4-chlorothiazole 8 is limited, 21,22 and here again, though we were able to reproduce one synthesis of this material in very low yield, 21 we found 8 was unstable as the free base and difficult to acylate, giving mixed products.
The acidity of the amide NH in thiazolides such as 1a and 1b suggested an alternative route to 4 0 -substituted thiazolides, viz.further acylation of N-protected versions of 7 and 8, followed by mild deprotection.We now report that t-butyl (4-oxo-4,5dihydrothiazol-2-yl)carbamate is an ideal, versatile precursor for such derivatives.Treatment of Boc derivative 6 20 with TFA in CH 2 Cl 2 , followed by basication with NaHCO 3 and extraction, afforded the free amine 7 in 94% yield, which proved rather unstable on storage and was used immediately, Scheme 2. Reaction of 7 with O-acetylsalicyloyl chloride 9 using two-phase acylation conditions 11,12 was quite unsuccessful.Instead, anhydrous acylation in THF using Et 3 N as base gave a slow, complex reaction.Workup aer 46 h at 20 °C gave two major products by chromatography, from which the desired thiazolide 10 was isolated in 17% yield and puried by recrystallisation.A major byproduct was apparently the acetamide 11 (m/z 221, 223) though this was difficult to purify fully.
Similarly, anhydrous acylation of 2-amino-4-chlorothiazole 8 21 with 9 again gave a slow complex reaction.By chromatography, the desired thiazolide 12 was obtained in 19% yield and further puried by recrystallisation.A signicant more polar product proved to be a bis-acylated derivative 13, which interestingly possessed a bis-acylamino rather than a tautomeric acylimino structure, as shown by single crystal X-ray analysis, Fig. 4. We therefore turned to alternative 2-aminothiazole intermediates.

Protected forms of pseudothiohydantoin
N-(4-Oxo-4,5-dihydrothiazol-2-yl)acetamide and its chlorination.Pseudothiohydantoin 14, sc.2-aminothiazol-4(5H)-one, is commercially available or easily prepared from thiourea and bromoacetic acid in a typical Hantzsch synthesis, 23 and carries built-in 4-substitution.As noted above, heating 14 with excess POCl 3 21 gave a very low yield of 2-amino-4-chlorothiazole 8.We therefore studied N-protected versions of 14, aiming rst at the acetamide, Scheme 3. Heating 14 with Ac 2 O/AcOH 24 led to a very slow reaction, even at 100-105 °C, so we switched to amine bases.Treatment of 14 with Ac 2 O and DMAP in THF at 20 °C gave a steady reaction and delivered very largely the previously unknown N, O-diacetate 15 in high yield; its structure was conrmed by a single crystal X-ray determination, Fig. 5, since other tautomeric products were possible.The use of Et 3 N gave a mixture of products including 15 and the desired monoacetamide.Use of the weaker base N-methylmorpholine at 60 °C gave a controlled reaction, which generated the desired acetamide 16 in very good yield with negligible diacetylation.Treatment of 16 with POCl 3 at 50 °C gave a very slow reaction until catalytic DMF was added; the 4-Cl compound 17 25a was then isolated in satisfactory yield.The same product was obtained in 72% yield by reaction of 16 with Ph 3 P and N-chlorosuccinimide (NCS) (1.5 eq.each; cf. next section) in MeCN at 20 °C.Another route claims chlorination of 2-acetamidothiazole using 'green' conditions, viz.NaCl and oxone, 25b but it is not clear whether the 4-Cl isomer 17 is the product since these authors' NMR data look signicantly different from ours.The reaction of 2-aminothiazole with 1-chloro-1,2-benziodoxol-3-one 22 was also stated to afford 17.
To seek reassurance on the regiochemical point, we studied the direct chlorination of 2-acetamidothiazole 18 with NCS in MeCN, Scheme 4. We used a similar procedure once before on a thiazolide. 11In fact this chlorination proceeded smoothly, using mild acid catalysis with Amberlyst A-15 (H + ) resin, and the product, isolated in unoptimised 65% yield, was shown to be the 5-Cl isomer 19 by a single crystal X-ray determination, Fig. 5.The 1 H and 13 C NMR data of this material were identical with those reported 25b and claimed to be the 4-Cl isomer.
Under relatively mild conditions (HCl, aq.MeOH, 50 °C) we found that hydrolysis of 17 gave rapid decomposition with reversion to 14.This probably resulted from ring protonation at C(5) followed by attack of water at C(4).
Tert-Butyl (4-oxo-4,5-dihydrothiazol-2-yl)carbamate and its halogenation.We therefore switched to Boc protection, to allow for mild anhydrous acidolysis eventually.Boc pseudothiohydantoin is disclosed in the patent literature, 27a prepared by reaction of di-t-butyl pyrocarbonate (Boc 2 O) with 14 in 15% yield using DMAP catalysis.Instead, using THF-water at pH 10 with Na 2 CO 3 or NaOH, a clean conversion to the mono-Boc derivative was obtained: 20 was isolated in 86% yield, Scheme 5.Under these conditions, formation of any bis-adduct is minimal and excess Boc 2 O is steadily hydrolysed.A later Pzer patent 27b cited a similar yield by heating 14 with two equivalents of Boc 2 O and no catalyst in THF at 60 °C for 48 h.
We anticipated that 20 would be readily enolised; hence reagents for the chlorination of other tautomeric hydroxy heterocycles such as 2-hydroxypyridine/2-pyridone under Appeltype conditions 28 should be effective.More recently, variants of the original Appel method using catalytic Ph 3 PO 29 and a sustainable procedure avoiding chlorinated solvents 30 have  For the introduction of Br at C(4), as noted earlier, the Br rearrangement ('halogen dance') 20a,b is feasible: the substrate (Scheme 1) is prepared from 2-amino-5-bromothiazole. 35 Here too, however, 20 proved a highly suitable intermediate, and on treatment with Ph 3 P and N-bromosuccinimide 33,34b 6 was readily obtained. 36Here the solvent choice was signicant, with MeCN denitely superior to CH 2 Cl 2 , giving 6 in 63% yield.Another good reagent proved to be ethyl tribromoacetate, 34b,37 again employing MeCN, which gave a virtually identical yield, though here purication was more difficult.It is noteworthy that MeCN oen proves a superior solvent in the Appel-type halogenation reaction 38 and may even divert the reaction to other products. 39Boc-amino-4-iodothiazole 22 was disclosed in a patent 40 as a useful intermediate for Suzuki couplings, but with no preparative detail.We obtained this compound in an unoptimised 28% yield by treatment of 20 with Ph 3 P and N-iodosuccinimide at 0-20 °C; a little free I 2 was used to initiate the reaction.41 In Scheme 6 we give a mechanism for these halogenations, using Cl 3 C$CN as the example donor, generating 21.
We also studied the reactions of both 16 and 20 with Middleton's DAST reagent, 42 hoping to gain access to 4-uoro derivatives: currently there is no reported preparation of

Acylation of Boc intermediates and thiazolide synthesis
The NH of compounds such as 21 is considerably more acidic than a typical amide 43 or even acetanilide (pK a = 13), 44  This sequence was equally applicable to the bromo intermediate 6, which via intermediate 24 gave 10, cf.Scheme 2. Clearly this sequence represents the method of choice for the synthesis of 10 and 12.

Conclusions
N-Boc protected forms of 2-amino-4-halothiazoles are readily available from Boc-pseudothiohydantoin, which is itself available from pseudothiohydantoin in high yield.The tendency of the heterocycle to exhibit tautomeric behaviour and to overreact with electrophiles is thus avoided.In general, N-halosuccinimides in conjunction with Ph 3 P under Appel-type conditions are effective reagents for the halogenation step, but Cl 3 CCN proved optimal for chlorination.Further acylation of these intermediates with O-acetylsalicyloyl chloride, followed by mild deprotection, offers high-yielding syntheses of 4-bromo and 4-chlorothiazolides.The relatively high acidity of amide NHs in derivatives such as 21 is signicant: this bis-acylation/ mild deprotection sequence may well offer good alternative syntheses for other heterocyclic amides.Direct acylation of the corresponding free amines 7 and 8, by contrast, gave low yields of mixed products.

General experimental procedures
Organic extracts were nally washed with saturated brine and dried over anhydrous Na 2 SO 4 prior to rotary evaporation at <30 °C.Moisture sensitive reactions were carried out in anhydrous organic solvents (purchased from Sigma-Aldrich) under a N 2 or Ar atmosphere.Reactions were monitored by analytical thinlayer chromatography using Merck Kieselgel 60 F 254 silica plates, and were viewed under UV or by staining with KMnO 4 or iodine.Preparative ash column chromatography was performed on either VWR Prolabo silica gel or Sigma-Aldrich silica gel (particle size 40-63 Å).Melting points were recorded using a Bibby-Sterlin Stuart SMP3 melting point apparatus and are uncorrected.Mass spectra were obtained in either electrospray mode (ES) with a Micromass LCT or chemical ionization (CI) mode with a Micromass Trio 1000 using ammonia.Elemental analyses were performed by Mrs Jean Ellis, University of Liverpool. 1 H and 13 C NMR spectra were obtained using a Bruker Avance or a Bruker DPX 400 instrument operating at 400 and 100 MHz, respectively; chemical shis are reported in ppm (d) relative to Me 4 Si.Coupling constants (J) are reported in Hz.

Fig. 5 26 Scheme 4
Fig. 5 Single crystal X-ray structures of 15 and 19.See ESI † for cif file data.The syn-orientation of the S atom and carbonyl oxygen in both cases results from nonbonding overlap between the C-S s* orbital and O lone pair electrons. 26
and our previous experience had indeed shown that further N-acylation was possible.Using Et 3 N as base, acylation of 21 with O-acetylsalicyloyl chloride cleanly afforded a 70% yield of the Boc intermediate 23 (Scheme 8).Mild acidolysis (dilute CF 3 CO 2 H, CH 2 Cl 2 ) then delivered thiazolide 12 in near quantitative yield, identical to the product obtained in low yield by acylation of 8, Scheme 2.